Mutation inyaaTLeads to Significant Inhibition of Phosphorelay during Sporulation inBacillus subtilis

Abstract

In the course of a Bacillus subtilis functional genomics project which involved screening for sporulation genes, we identified an open reading frame, yaaT, whose disruptant exhibits a sporulation defect. Twenty-four hours after the initiation of sporulation, most cells of the yaaT mutant exhibited stage 0 of sporulation, indicating that the yaaT mutation blocks sporulation at an early stage. Furthermore, the mutation in yaaT led to a significant decrease in transcription from a promoter controlled by Spo0A, a key response regulator required for the initiation of sporulation. However, neither the level of transcription of spo0A, the activity of H , which transcribes spo0A, nor the amount of Spo0A protein was severely affected by the mutation in yaaT. Bypassing the phosphorelay by introducing an spo0A mutation (sof-1) into the yaaT mutant suppressed the sporulation defect, suggesting that the yaaT mutation interferes with the phosphorelay process comprising Spo0F, Spo0B, and histidine kinases. We also observed that mutation of spo0E, which encodes the phosphatase that dephosphorylates Spo0A-P, suppressed the sporulation defect in the yaaT mutant. These results strongly suggest that yaaT plays a significant role in the transduction of signals to the phosphorelay for initiation of sporulation. Micrographs indicated that YaaT-green fluorescent protein localizes to the peripheral membrane, as well as to the septum, during sporulation.Initiation of sporulation in Bacillus subtilis is regulated by a signal transduction pathway, the phosphorelay, which is a multicomponent phosphotransfer system that is switched on in response to environmental, cell cycle, and metabolic signals (3, 36). The processing and integration of these signals by the phosphorelay control the level of phosphorylation of the transcription factor, Spo0A. Environmental and cellular signals that favor sporulation activate autophosphorylation of the sensor kinases KinA, KinB, KinC, and KinD, leading to input of a phosphate group into the phosphorelay (1,2,12,13,18,20,33,37,46). In this relay, the phosphate group is subsequently transferred to a response regulator, Spo0F. The resulting molecule, Spo0F-P, serves as a substrate for the Spo0B protein, a phosphotransferase which finally activates Spo0A by transferring the phosphate to the Spo0A protein (2). Spo0A-P, the activated form of Spo0A, indirectly controls the transcription of a number of genes by regulating the level of other transcription regulators. Spo0A-P directly activates transcription of the genes for many regulatory proteins and sigma factors required for cell-type-specific gene expression. Spo0A-P is also known to stimulate axial filament formation and asymmetric polar septation, which give rise to two unequal cells, a larger mother cell and a forespore cell (21, 38).Just after septation, gene expression is controlled by the RNA polymerase sigma factors, F in the forespore and E in the mother cell. Later in sporulation, when the forespore has become engulfed by the mother cell, F and E are r...

The RicAFT (YmcA‐YlbF‐YaaT) complex carries two [4Fe‐4S]2+ clusters and may respond to redox changes

“…Early work showing a role for the RicAFT complex in Spo0A‐dependent transcriptional regulation suggested a role for the complex in control of the phosphorelay. This conclusion has been supported by both biochemistry and genetic suppressor analyses (Tortosa et al ., ; Hosoya et al ., ; Kearns et al ., ; Carabetta et al ., ; Dubnau et al ., ). In particular, the direct stimulation of the phosphorelay in vitro supports the idea that this is an important function of the complex.…”

“…Past work has shown an effect of individual yaaT , ylbF and ymcA null mutations on Spo0A∼P‐dependent transcription (Tortosa et al ., ; Hosoya et al ., ; Branda et al ., ; Kearns et al ., ). The sof‐1 allele of spo0A that bypasses the phosphorelay by permitting the direct phosphorylation of Spo0A, can bypass the requirement of yaaT for sporulation, as well as spoIIG transcription (Hosoya et al ., ; Dubnau et al ., ). Since spoIIG transcription only requires Spo0A∼P these findings demonstrate that the complex of YlbF, YmcA and YaaT is required for increased phosphorelay activity.…”

The Cell Wall Regulator σISpecifically Suppresses the Lethal Phenotype ofmblMutants inBacillus subtilis

“…However, the ylxA deletion proved to be not very potent in suppressing the Mg 2ϩ dependency of the mbl deletion strain (data not shown). One transposon insertion each was found in yaaT encoding a protein involved in the phosphorelay cascade during initiation of sporulation (29), in the gene for the glutamate transporter GltT (51,54), and in pnpA which codes for polynucleotide phosphorylase (39,43,59). We focus in the present study on the most potent suppressor, generated by insertion in rsgI.…”

A complex of YlbF, YmcA and YaaT regulates sporulation, competence and biofilm formation by accelerating the phosphorylation of Spo0A

“…The overall goal of this work was to clarify the mechanism by which YmcA and YlbF function in development. To this end, we used a proteomics approach to identify novel interacting partners of YmcA and YlbF and discovered that both interact with a third protein, YaaT, which had previously been shown to act early during sporulation (Hosoya et al ., ). YmcA, YlbF and YaaT interact directly with one another, forming a stable ternary complex in vitro .…”